Introduction

"The nutrients are important and it’s important to be flexible and be able to change according to what is available on the market. This sounds easier than it is. Manufacturing is about up-scaling and it’s one of the challenges in synthetic biology."

Gernot J. Abel, Research and Technology, Novozymes

We are from a technical university in a small country where innovation is in focus, so have been encouraged to think outside the box throughout our entire studies. Although, since we are just a group of young, blue eyed students coming from the comfort of the university, we learned a lot by investigating how our idea fit into real life settings and exploring the viability of basing industrial biotechnological processes on waste streams.

Throughout our project, from the initial development of the idea to finalizing the lab work, we have been in close contact and collaboration with the industry. We started out searching for abundant waste streams in Denmark by contacting and visiting local factories. Afterwards we presented our idea to players in the biotech manufacturing companies, and the feedback we recieved from them made us more aware of the challenges our technology is facing and it ultimately led us to a more feasible project.

Investigating local industrial waste streams

We started investigating one of the classical examples of a waste stream today, glycerol, and came in contact with local biodiesel producers, Perstorp and Emmelev A/S. We investigated their waste streams with the perception that glycerol was an abundant waste product from their productions and thereby a by-product, they would be more than happy to get of their hands. Talking to the Biodiesel producers, we were suprised to find that they do not percieve glycerol as being a waste, but actually already is a valuable byproduct used in the chemical, pharmaceutical and tobacco industry. After realizing this we went back to the drawing board in search of a "true waste product", which did not have any competing uses. After researching less classical waste streams and talking to even more industry, we got into contact with a local canola oil producer who had a promissing substrate for our project. His process of producing cold pressed canola oil produced around 1 - 1.6% of a lipid- and carbohydrate rich sediment, which he could not recycle in the process or use in animal feed, due to its high content of polyols. Instead the sediment was more an annoyance for him, as he was forced to burn it to dispose of it. He was excited about our project, as this would allow his by-product to be transformed into something useful for society. Talking to him we learned a lot about canola oil production and waste handling, and quickly realised that this by-product was ideal for our project. We integrated this in our project by investigating the growth characterstics of Y. lipolytica on the sediment and finally settling for it as a proof of concept substrate for the biorefineries of tomorrow.

×

DESCRIPTION

Implementation of substrates from waste streams in the biotech manufacturing industry

The second part of our integrated human practices consisted of interviews and feedback from two of the biggest biotech companies in Denmark, Novo Nordisk A/S and Novozymes A/S, who we asked questions about our idea about using complex waste streams, such as the sediment from canola oil production, in state of the art industrial processes.

Interview with Novo Nordisk A/S

Novo Nordisk develops and produces more than 50% of the insulin worldwide along with growth factors and medicine for hemophilia. To get the perspective of Novo Nordisk on how the increasing demand for insulin and higher demand of crops/edible carbon sources will require a more sustainable technology, we went to visit the Novo Nordisk Science Park in Måløv, Denmark, to interview Nina Gunnarsson and Kjeld Olesen, who both are researchers in the Strain Development department in Novo Nordisk. With more than five years of employment at the company, they provided us with some insights on how they currently produce insulin, what they value in a production organism and what it would take for them to apply our technology. Click here to read the full interview or read along to get a short version.

Novo Nordisk currently produces recombinant insulin in the conventional baker's yeast, Saccharomyces cerevisiae, by growing it on different refined sugars.

“We use a very little amount of sugars compared to what is consumed for food, so right now that sort of sustainability is not that a big of a concern for us. We are more focused on our obligation to provide sufficient amounts of insulin to the patients”

said Kjeld Olesen, when we asked them if changing their substrate into a less competitive and sustainable one was something they were considering. Nina Gunnarsson added: “We obviously have a big focus on having the most optimal processes with the highest possible yield with the minimum environmental impact.” And the most optimal process is for Novo Nordisk is one, where the demand for purity is met. Nina and Kjeld explained to us that purity in regards to avoiding degradative proteins and post-translational modification, especially O-glycosylation, is very important, but the purity of the entire process is essential to obtain the high level of quality required for a therapeutic protein.

We ultimately asked Nina and Kjeld from Novo Nordisk, if they believed changing their insulin production to Yarrowia lipolytica growing on abundant waste streams would be an option: “Well, it would be a huge project! Because the product quality is so important, it would be a very long and complicated changeover, which will include a new factory and new experts, so it would have to be a very good business case,” Nina replied and Kjeld followed: “A stabile, continued supply of substrates is very important. The content of the substrate cannot vary either.” And in conclusion, Nina suggested we should consider applying our technology for something other than therapeutic compounds: “Maybe this would be more applicable for production of other things than therapeutics - such as bulk chemicals, enzymes or food additives, that do not have the same quality and purity requirements.”

Summary of the main points from the Novo Nordisk interview:

• Purity and quality of the product along with a stable supply chain and production is the most important aspects when producing therapeutics.
• Triple bottom line mindset (economic, social and environmental sustainability) and a strong focus on the obligation of meeting the insulin demand.
• Novo Nordisk sees our technology as more applicable for production of less regulated compounds such as bulk chemicals, enzymes or food additives.

×

DESCRIPTION

Interview with Novozymes

With this in mind, we turned to another major league player in the biotechnological industry - Novozymes A/S. Novozymes has a very diverse production line and produces enzymes for industries ranging from agriculture, household care and food to bioenergy and wastewater solutions. Furthermore, they are a top 10 company on the Fortune Magazine’s Change the World list, so we thought - who else would be more likely to seize our idea and technology? To read the full interview, you can click here.

“I’m not saying that it IS a challenge, but it could be a challenge. If you cannot get the dirt out of the fermentor, then you need to use organic solvents and it might also have other consequences. But in general, I would say yes! There is a lot of available fat sources that could be used. But we need to know the consequences.” was what Gernot J. Abel from the Research and Technology department at Novozymes in Bagsværd, Denmark, said, when we asked him, whether Novozymes would consider switching to Y. lipolytica as production organism in case our molecular toolbox is up and running, and then base the production on waste streams such as glycerols, fatty acids and media containing other complex carbohydrates. “The more you can do with one single strain, the better. But if Yarrowia is excellent making more homogeneous proteins, then that would be a selling point. Get rid of your waste, so less waste and more value. If you can cut production costs 10% down and use different waste streams, then cool!”

Novozymes currently base their production on easy accessible proteins and carbohydrates such as sugars and starch and as cheap as possible, but Gernot J. Abel is open to the idea of using waste streams as substrates. However, he also made us aware of some the main points of the Novozymes strategy, which our technology must accommodate in order to be viable. Gernot stated that having a process, where the fermentor is cleanable and the fermentation controllable is essential: “One question is if we can make these cell factories cheaper… It has to be controlled, it needs to be clean and temperature controllable. You have to bring in all the nutrients. Cell factories is our heart and our production system, so it’s extremely important.”

Like Novo Nordisk, Novozymes have one main goal and that is to deliver, what the customers require both in regards to meeting the demand for quality and quantity.

“Novozymes is about manufacturing. Our RD research is important in developing but in the end of the day, you have to deliver to the market and be able to send enzymes and ship them to the customers... For us it’s more important that our enzymes can do their job. We keep trying to optimize our enzymes to perform better. That is where our focus is compared to changing our production host.”

Summary of the main points from the Novozymes interview:

• The process should be cleanable and controllable.
• Production on waste streams should be cheaper compared to traditional production.
• Quality and quantity of the product is the most important.

×

DESCRIPTION

Exploring the possibilities

After the interviews with Novo Nordisk and Novozymes, we were even more confident in our project. Insulin is most likely not produced from waste sources anytime soon, although the tools we have developed during this project might actually yield a viable production of bulk products in the not so distant future.